Ranibizumab ELISA Assay Kit
The Ranibizumab ELISA Assay Kit is For Research Use Only
Size: 12×8 wells
Sensitivity: <1.25 ng/mL
Standard Range: 1.25-80 ng/ml
Incubation Time: 5 hours
Sample Type: Serum, Plasma, Cell Culture Supernatants
Sample Size: 100 µL
Alternative Name: Lucentis
The method employs the competitive enzyme immunoassay technique. Antibodies to Ranibizumab are precoated onto microwells. Samples or Standards and Ranibizumab-Biotin Conjugate are pipetted into microwells and human Ranibizumab present in the sample or standard will compete with Ranibizumab-Biotin Conjugate and bound by the capture antibody. After washing microwells, HRP conjugate is pippeted and incubated. Free HRP conjugate will be removed by washing cycle.The ready to use substrate solution (TMB) is added to microwells and color develops inversely proportional to the amount of Ranibizumab in the sample. Color development is then stopped by addition of stop solution. Absorbance is measured at 450 nm.
Sample Preparation and Storage:
Blood is taken by venipuncture. Serum is separated after clotting by centrifugation. Plasma can be used, too. Lipaemic, hemolytic or contaminated samples should not be run. Repeated freezing and thawing should be avoided. If samples are to be used for several assays, initially aliquot samples and keep at – 20°C. For Cell Culture Supernatant – If necessary, centrifuge to remove debris prior to analysis. Samples can be stored at -20°C or -80⁰C. Avoid repeated freeze-thaw cycles.
Ranibizumab mAb-based ELISA Assay
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Alemtuzumab ELISA Assay Kit
Ranibizumab (Lucentis) is a recombinant humanized IgG1 monoclonal antibody fragment that binds to and inhibits vascular endothelial growth factor A (VEGF-A). VEGF is a biochemical signal protein that promotes angiogenesis throughout the body and in the eye. Through binding to VEGF-A, ranibizumab interrupts the interaction of VEGF with its receptors, and thus prevents the subsequent growth of new blood vessels.
1. It is strongly recommended that all Controls and Samples be run in duplicates or triplicates. A standard curve is required for each assay. All steps must be performed at 37°C
2. Add 100 μl of Standards or Samples into the respective wells.
3. Cover the plate and incubate for 120 minutes at 37°C.
4. Pipette 50 μl of Ranibizumab: Biotin Conjugate into each well.
5. Cover the plate and incubate for 60 minutes at 37°C
6. Aspirate and wash plate 4 times with Wash Buffer (1X) and blot residual buffer by firmly tapping plate upside down on absorbent paper. Wipe off any liquid from the bottom outside of the microtiter wells as any residue can interfere in the reading step.
7. Add 100 μl of Streptavidin: HRP Conjugate into each well.
8. Aspirate and wash plate 4 times with Wash Buffer (1X) and blot residual buffer by firmly tapping plate upside down on absorbent paper. Wipe off any liquid from the bottom outside of the microtiter wells as any residue can interfere in the reading step.
9. Add 100 μl of TMB Substrate in each well.
10. Incubate the plate at 37°C for 60 minutes in dark. DO NOT SHAKE or else it may result in higher backgrounds and worse precision. Positive wells should turn bluish in color.
11. Pipette out 100 μl of Stop Solution. Wells should turn from blue to yellow in color.
12. Read the absorbance at 450 nm with a microplate reader.